The goal of this research program is to identify the CNS mechanisms that are responsible for integrating cardiovascular control with ongoing metabolic and behavioral needs. The parabrachial nucleus (PB), a major integrative center that links the forebrain and medullo-spinal cardiovascular control areas, plays a key role in this process. Two PB subdivisions, the external lateral (PBel) and dorsal lateral (PBdl) subnuclei, are intense terminal fields for ascending projections from the cardiovascular portion of the NTS. Electrical and chemical stimulation in the PBel produces a tachycardic/pressor/tachypneic response; PBel projects primarily to the amygdala and basal forebrain. stimulation in the PBdl produces a bradycardic/depressor response; its terminal fields are mainly in the hypothalamus. This project will focus on identifying the chemical neuroanatomy of the afferent and efferent connection of these two cell groups. In the first series of experiments, injections of retrograde tracers will be made into the PBdl and the PBel, and combined with immunohistochemical staining for putative neuromodulators in their afferent projections. The co-localization of two or more putative modulators in the same afferent neurons will also be studied. This information will be used to place injections of the anterograde tracer, PHA-L into these afferent cell groups. Individual afferent axons will also be stained using an immunofluorescence method for putative neuromodulators. The second series of studies will examine the collateralization of axons that provide these afferent inputs, by simultaneously injecting fluorescent dyes into either the PBdl or the PBel, and staining the retrogradely labeled neurons immunohistochemically to identify their content of putative neuromodulators. The third series of experiments will examine the chemical specificity of the PBel and the PBdl efferent connections by placing injection of PHA-L into these cell groups and simultaneously staining individual axons for neuromodulator immunoreactivity. Based on these results, a complementary series of experiments will be done, in which injections of retrograde fluorescent tracers will be placed into the Pbel and the PBdl terminal fields, and the retrogradely labeled neurons stained for neuromodulator immunoreactivities. Studies will also be done to examine the co-localization of neuromodulator immunoreactivities in these efferent neurons. Finally, in the fourth series of studies, the collateralization of the efferent projections from the PBel and the PBdl will be studied, using injections of different retrograde fluorescent tracers into pairs of terminal fields. These experiments will provide detailed information on the input-output relationships of the PBel and the PBdl, and their chemical specificity. this information will be critical for the design of future physiological and pharmacological strategies for investigating the role of the PB in central cardiovascular control.